Figure 1. The expression of G protein subtypes in the efferent ductules and ADGRG2 promoter-labeled non-ciliated cells.

(A) qRT-PCR analysis of mRNA transcription profiles of G proteins in brain tissues and the efferent ductules of WT (n = 3) male mice. Expression levels were normalized to GAPDH levels. *p<0.05, **p<0.01, ***p<0.001, efferent ductules compared with brain tissue. (B) Co-localization analysis of ADGRG2 (red fluorescence) and acetylated-tubulin (green fluorescence) in the efferent ductules of WT mice. Scale bars, 50 μm. (C) Co-localization of ADGRG2 (green fluorescence) and RFP (red fluorescence) in the same cells of male murine efferent ductules infected with the ADGRG2 promoter RFP adenovirus in WT mice. Scale bars, 50 μm. (D) qRT-PCR analysis of mRNA transcription profiles of G protein subtypes in brain tissues and isolated ADGRG2 promoter-labeled non-ciliated cells derived from the efferent ductules of WT (n = 3) male mice. Expression levels were normalized to GAPDH levels. *p<0.05, **p<0.01, ***p<0.001, ADGRG2 promoter-labeled efferent ductule cells compared with brain tissues. n.s., no significant difference. At least three independent biological replicates were performed for Figure 1A and D.

Figure 1—figure supplement 1. ADGRG2 is specifically expressed in non-ciliated cells.

(A) Control experiments: Direct immunofluorescence staining of secondary antibodies used in the manuscript (including donkey anti-sheep, red fluorescence; and donkey anti-rabbit, green fluorescence) in WT male mice efferent ductules. Scale bars, 50 μm. (B) Microscope analysis of efferent ductules and the nucleus in WT male mice, including a light image. Scale bars, 50 μm. (C) Bar graph representation and statistical analyses of co-localization of ADGRG2 and acetylated-tubulin in WT male mice efferent ductules (corresponding to Figure 1B in the main manuscript), n = 3 mice per group; 4–10 random areas were selected from each section, and six sections were randomly selected from each mouse.

Figure 1—figure supplement 2. The construction of the mouse ADGRG2-promoter-RFP used in the labeling of ADGRG2-expressed cells.

(A–B) Schematic representation of the construction of the mouse ADGRG2-promoter-RFP used in the labeling of ADGRG2 expressed cells in the epididymal efferent duct epithelium. Sub-cloning strategy of the ADGRG2-promoter (A). Schematic diagram of ADGRG2-promoter-RFP adenovirus vector (B). (C) Isolated epididymal efferent duct epithelium infected with the ADGRG2-promoter RFP adenovirus specifically labeled the ADGRG2-expressing non-ciliated cells. Scale bars, 50 μm.